Granulocyte colony-stimulating factor (G-CSF) is an important hematopoietic growth factor belonging to a larger class of biologically relevant glycoprotein cytokines. Two therapeutic forms of G-CSF are used clinically for the treatment of neutropenia induced by chemotherapy. The principal difference between these forms is the presence or absence of an O-linked carbohydrate at threonine-133. An established platform to prepare both therapeutic forms by means of chemical synthesis could address concerns with regard to the importance of G-CSF glycosylation. Additionally, further development of this research strategy would allow for modular access to more efficacious variants of G-CSF. For these reasons, glycoprotein G-CSF is considered a target for chemical synthesis. The development of efficient synthetic methods for the preparation of such therapeutic glycoproteins is an ongoing goal in our laboratory. This proposal entails the investigation of reaction scope and limitations with regard to our recently described solid-phase fragment coupling (SPFC) method. This method will be useful for the convergent preparation of large polypeptides and glycopeptides by the coupling of C-terminal polypeptide thioacids and N-terminal polypeptides on solid-phase. The application of SPFC technology would expedite the synthesis of target glycoprotein, G-CSF. To advance this method, the following synthetic challenges will be addressed: (1) the efficient preparation of C-terminal thioacids (+15-mers) and (2) the isonitrile mediated fragment coupling of C-terminal thioacids with resin-bound N-terminal polypeptides to access large polypeptides (+30-mers). Following this advance, the utility of SPFC technology will be examined in the context of target polypeptide fragments relevant to G-CSF synthesis. These targeted polypeptides will be assembled by chemical ligation techniques to prepare homogenously glycosylated and non-glycosylated forms of G-CSF.